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Biomedical subjects

A G Ewing

Publications and source records attributed to A G Ewing.

18 recordsLinked to original sources
PubMed1992

Characterization of glucose microsensors for intracellular measurements.

Ultrasmall glucose sensors have been constructed by using platinum-deposited carbon ring microelectrodes with glucose oxidase. Response times as low as 270 ms have been obtained with these sensors. Moreover, there is a linear relationship between sensor tip diameter and response times. The use of these sensors has been demonstrated in the detection of glucose in single-cell cytoplasm of the large dopamine cell of the pond snail Planorbis corneus. Current responses obtained at these sensors implanted into a cell increase following injection of 2 pL of glucose solution (3 M) into the cell. Results obtained from these experiments show that these sensors are suitable for glucose monitoring in ultrasmall environments. In addition, characterizations of these sensors have been investigated under different O2 concentrations. At atmospheric oxygen concentrations, glucose levels in the submillimolar range can be measured without oxygen interference; however, oxygen interference can be substantial at low oxygen concentrations.

Animals↗
PubMed1992

Voltammetric measurement of oxygen in single neurons using platinized carbon ring electrodes.

Naflon-coated ultrasmall platinum ring electrodes have been implanted in the giant dopamine neuron of the pond snail Planorbis corneus, and the oxygen concentration inside these single neurons has been estimated. Experimental data suggest that the intracellular oxygen level in the identified dopamine neuron of P. corneus is approximately 0.032 mM. The oxygen concentration immediately outside the cell (ca. 10 microns away from the cell) is 0.041 mM. Furthermore, staircase voltammetry can be used to monitor dynamic changes in oxygen concentration inside the cell after bathing with Ringer's solution saturated with air/oxygen. Data obtained for intracellular oxygen concentrations suggest that intracellular oxygen consumption is increased following potassium chloride-induced stimulation of these cells.

Animals↗
PubMed1992

Electroosmotic flow control and monitoring with an applied radial voltage for capillary zone electrophoresis.

The control of electroosmotic flow by electronic means for capillary zone electrophoresis is presented. This is accomplished by the application of a radial voltage field with a rugged and flexible conductive polymer sheath. Fundamental theory for effect of the applied radial voltage on electroosmotic flow is developed. In addition, the effects of surface pretreatments are examined and compared to trends predicted by the theory.

Electric Conductivity↗
PubMed1990

Capillary electrophoresis in 2 and 5 microns diameter capillaries: application to cytoplasmic analysis.

Capillary electrophoresis with electrochemical detection in 2- and 5-micron capillaries has been developed to study ultrasmall biological environments. Sample volumes as low as 270 fL have been injected into the electrophoresis capillary with subattomole detection limits for easily oxidized species. We have applied this method to the analysis of single cell cytoplasm. Sampling of the cytoplasm is accomplished by inserting one end of the electrophoresis capillary directly into a single nerve cell. The high-voltage end of the electrophoresis capillary has been etched with hydrofluoric acid to form a microinjector. This injection scheme represents an improvement over those previously used for similar applications. The excellent selectivity of this method is demonstrated for catecholamine and indolamine neurotransmitters and their metabolites found in the invertebrate system, the pond snail Planorbis corneus.

Animals↗
PubMed1990

Capillary electrophoresis with indirect amperometric detection.

The use of indirect amperometric detection with capillary electrophoresis is demonstrated. The system consists of a porous glass coupler which allows amperometric detection at a carbon fiber electrode placed in the end of the capillary. 3,4-Dihydroxybenzylamine is added to the buffer system as a continuously eluting electrophore. Indirect amperometric detection in 9-mumol I.D. capillaries provides detection limits as low as 380 attomole for the amino acid arginine. Finally, both direct and indirect amperometric detection can be accomplished simultaneously.

Amino Acids↗
PubMed1990

Dopamine concentration in the cytoplasmic compartment of single neurons determined by capillary electrophoresis.

It has long been a goal of the biochemist to study the chemistry of single cells. This is particularly important in neuroscience, since each cell can maintain a specific function and identity. Investigations into neurotransmitter compartmentalization within the cell body of neurons should lead to an understanding of the role of the cell body in metabolism, uptake and storage of neurotransmitters. Using capillary electrophoresis, it has been possible to separate and detect attomole levels of neurotransmitter in picoliter volumes of cytoplasm withdrawn from single neurons of the pond snail, Planorbis corneus. This work demonstrates, for the first time, the direct determination of the cytoplasmic concentration of dopamine in single, intact neurons.

Cytoplasm↗
PubMed1990

Estimation of free dopamine in the cytoplasm of the giant dopamine cell of Planorbis corneus by voltammetry and capillary electrophoresis.

Voltammetric electrodes having a tip diameter of 2-12 microns and microscale capillary zone electrophoresis have been used for dynamic and static monitoring of dopamine in the somal cytoplasm of the giant dopamine neuron of Planorbis corneus. Intracellular dopamine levels can be altered by extracellular application of dopamine or ethanol. The latter experiment provides a means to estimate total stores of endogenous dopamine, and the evidence presented suggests that at least 98% of these stores are bound and not directly accessible to the cytoplasm.

Animals↗
PubMed1989

Separation of serotonin from catechols by capillary zone electrophoresis with electrochemical detection.

Capillary zone electrophoresis with electrochemical detection is demonstrated with columns having only 9-microns inner diameter. Amperometric detection limits of 0.7 amol are reported for serotonin. The difficult problem of resolving serotonin and dopamine--two neurotransmitters of interest having similar electrophoretic mobilities--is addressed by chemical means to improve selectivity. These include buffer modification with 2-propanol and a system employing borate complexation of the catechol in combination with sodium dodecyl sulfate micelles.

Catecholamines↗
PubMed1989

Capillary electrophoresis.

The combined techniques that comprise capillary electrophoresis are under rapid development for a wide range of molecules and sample types. The capillary format provides fast, highly efficient separations and makes automation possible. At present, systematic approaches to separation methodology are still in the development stages and better detection schemes need to be developed. There is no doubt that electrophoresis in a capillary tube will take its place as a substitute for HPLC in specific applications. This is especially apparent when analysis time, efficiency, or simplicity is of greatest concern. Capillary electrophoresis will prove extremely useful for separations of nucleotides and proteins for analytical purposes in biotechnology. Finally, the ability to use ultrasmall-volume samples makes capillary electrophoresis ideal for the development of separations-based sensors for the analysis of microenvironments.

Electrophoresis↗
PubMed1988

Retention of ionic and non-ionic catechols in capillary zone electrophoresis with micellar solutions.

The use of micellar solutions in capillary zone electrophoresis has been primarily relegated to separations of non-ionic solutes, while its applicability to cationic species has been unexplored. We have found that the use of sodium dodecyl sulfate micelles in phosphate buffer allows for tremendous gains in selectivity for several cationic and non-ionic catechols over what can be obtained with normal capillary zone electrophoresis. Complexation of catechols with boric acid alters the net charge on the solutes and changes the partitioning behavior to produce adequate selectivity with improved analysis times. Although the mechanisms of solute interaction with the micellar phase for the cationic species are not decisively known, evidence is presented supporting the existence of ion-pairing equilibria simultaneously accompanied by micellar solubilization.

Borates↗
PubMed1985

Amphetamine attenuates the stimulated release of dopamine in vivo.

Carbon-fiber voltammetric electrodes have been used to measure the release of dopamine in the caudate nucleus of an anesthetized rat. Release is induced by electrical stimulation of the medial forebrain bundle. The amplitude of the observed release is attenuated by i.p. injection of amphetamine. A similar attenuation is induced by reserpine; however, at a slower rate. The combined regimen of amphetamine (1 or 10 mg/kg) and electrical stimulation does not deplete striatal dopamine levels and thus the decreased release of dopamine is not a result of depleted dopamine stores. Benztropine (25 mg kg-1) is able to cause a short term inhibition of the action of amphetamine (1 mg kg-1). The dopamine agonist pergolide (0.5 mg kg-1) does not affect the stimulated release. Haloperidol (1.0 mg kg-1) increases the amount of DA release, but is unable to attenuate the inhibition caused by amphetamine. Thus, it appears that the actions induced by amphetamine are a result of interaction with the neuronal uptake carrier and subsequent transport of dopamine from a functional to nonfunctional pool. In isolated striatal synaptic vesicles, amphetamine is found to block dopamine uptake and induce its release. This in vitro evidence provides a possible mechanism for the observed in vivo actions of amphetamine.

Animals↗